In this proposal Dr. Starr will use Sleeping Beauty (SB) transposon-based mutagenesis to screen for novel cancer genes. The protein products of identified genes will be analyzed for their role in human cancer by altering their levels in cell lines and measuring the effect on proliferation, apoptosis, colony formation, and signaling pathways. The SB system has been used to drive tumorigenesis in mice and subsequently identify both known and novel genetic mutations that drive leukemias, sarcomas, hepatocellular carcinomas, and colorectal cancers. Some tissue types, however, have proven refractory to SB mutagenesis. To successfully screen for genes in these tissues Dr. Starr has proposed several modifications to the SB system. Dr. Starr has been instrumental in developing the SB system during his tenure as a post-doctoral fellow in the Masonic Cancer Center (MCC) at the University of Minnesota. To help Dr. Starr transition to an independent research career, he has assembled an advisory committee consisting of experts in several different areas critical for the success of his research and career development. In addition to the direct support provided by his Mentor and advisory committee, Dr. Starr will use the shared resources available in the MCC. These include core facilities in biological imaging, tissue procurement, flow cytometry, comparative pathology, cytogenetics, bioinformatics/biostatistics, RNA interference, high-throughput sequencing, and a Mouse Genetics Laboratory capable of generating transgenic, knock-out, and knock-in mice. The candidate intends to continue his post-doctoral fellowship for two years while working on Aims 1 and 2 of this research proposal. Dr. Starr's long-term goal is to secure a position as an independent investigator in an academic setting where he plans on dedicating his career to understanding the genetic mechanisms driving cancer. Studies documenting genetic changes within human tumor genomes confirm that a large number of genetic defects contribute to tumor growth, yet only a small subset have been characterized. Furthermore, within a single tumor type there is great heterogeneity between individual tumors. The studies in this proposal address a critical need in the field of cancer research to develop better methods of identifying the subset of genetic changes that are actually driving tumor growth. In Specific Aims 1 and 2, several novel candidate cancer-driving genes identified by Dr. Starr in a screen for colorectal cancer (Wac, Tcf12, and Rspo2) and lung cancer (Akap13, Cul3, and Map4k3) will be studied to determine their specific role in human cancer. The levels of these proteins will be altered in human cell lines, both normal and transformed, to determine their effect on growth, apoptosis, the ability to form colonies in soft agar, and regulation of key signaling pathways. Aim 3 proposes several improvements to the SB system, using newly developed highly active transposases and transposons, to uncover novel lung cancer genes. The results of these studies can be used to develop a more personalized approach to cancer therapy. PUBLIC HEALTH RELEVANCE: The goal of this project is to use an animal model to understand the genetic basis of lung and colorectal cancer. The genetic mutations discovered using this animal model will then be functionally tested in human lung and colorectal cancer tissue and cell lines. The results of these studies will help us decide how to better treat these common, deadly cancers.